Microtubule sliding and dynein mechanochemistry are the basis for motion in several biological systems, particularly motion that occurs in cilia, flagella and spermtails, and apparently the mitotic apparatus. As a model system, Triton X-100 demembranated Tetrahymena cilia can be caused to actively disintegrate or slide apart upon reactivation with MgATP2- which is the primary substrate for the dynein ATPase that forms transient crossbridges in order to produce active sliding. The dynein crossbridges can be stabilized in an attached state by incubation in 1-3mM MgSO4. By using the disintegration reaction, we wish to determine the various structural conformations of the dynein arms as related to active sliding and substrate conditions. We also are concerned with the physiological nature of the Mg ions dependent crossbridge reaction. Does the stable attached state manifest part of the crossbridge cycle? This question will be studied by analyzing the sensitivity of the attached crossbridges to ATP, non-hydrolyzable nucleotide analogues and different divalent cations. Similarly, the reaction or recombination of purified 30S (whole arm) dynein with both A- and B-subfiber sites will be studied in terms of its cation dependence and nucleotide sensitivity. Since the Mg ions dependent crossbridge reaction is systematically sensitive to ATP, we also will study the relationship of the resulting crossbridge frequencies and distributions to the active beat cycle of Unio gill cilia. In Tetrahymena cilia potential regulatory cations such as Ca ions do not appear to have a direct effect on dynein crossbridging activity and hence active sliding. However, Ca ions does increase the sensitivity of the attached crossbridges to ATP by several times. Is this increased sensitivity a manifestation of possible regulatory activity by the cation? We will analyse the effects of CA ions on the crossbridge reaction, particularly as those effects are related to crossbridging frequencies and active sliding. These studies can be expected to yield direct information of the physical nature of dynein crossbridging activity and microtubule sliding, and hopefully identify one or more states of the mechanochemical reaction sequence.